PrP is a normal endogenous protein of unknown function in brain and a variety of other mammalian tissues. In the brains of animals afflicted with scrapie and other transmissible degenerative neuropathies, PrP accumulates in a proteinase K (PK)-resistant form which can aggregate into fibrils and form amyloid-like plaques. There is substantial evidence that the formation of the PK-resistant PrP is scrapie specific and important in the pathogenesis of the disease. However, it is not yet clear whether the PK-resistant is the transmissible agent itself, a component of the agent, or a pathological byproduct of the disease. Since the replication or pathogenesis of the scrapie agent may involve the production of PK-resistant PrP, we have continued studies of the properties and biosynthesis of both the PK-resistant and normal forms of PrP in tissue culture cells. We have determined that in scrapie-infected neuroblastoma cells, the two forms of PrP dimer in their aggregation states, N-termini, turnover rates and sensitivities to protease and phospholipase treatments of intact cells. We have extended our studies of normal PrP biosynthesis to rat PC12 cells, and have found that the size of PrP precursors and products, the kinetics of PrP biosynthesis, and the linkage of PrP to the plasma membrane by phosphatidyl inositol are similar to those in mouse neuroblastoma cells. Studies of the scrapie agent derived from mouse neuroblastoma cells have indicated that, like the brain-derived agent, ft can be slowly neutralized by high concentrations of PK. This indicates that has an essential protein component. However the infectivity is resistant to treatment with a variety of nucleases suggesting that it contains no nucleic acid or that the nucleic acid is protected by other components of the preparation. It is possible that a conformational abnormality accounts for the characteristics of PK-resistant PrP. To investigate this possibility, we have purified PK-resistant PrP from scrapie brain tissue and are initiating secondary structure analysis of it by infrared spectroscopy. Ultimately, we hope to be able to ascertain whether its conformation differs from that of the normal PrP.